IDENTIFICATION OF ENDOPHYTIC BACTERIA ASSOCIATED WITH N2 FIXATION AND INDOLE ACETIC ACID SYNTHESIS AS GROWTH PROMOTERS IN CURCUMA ALISMATIFOLIA GAGNEP.

Endophytic bacteria carrying out dinitrogen (N₂) fixation and indole acetic acid (IAA) synthesis were firstly identified in C. alismatifolia, a globally important flower crop. Their potential as growth promoters to stimulate the rapid growth of host plant was also examined. It will be beneficial to reduce the propagation period of tissue culture plantlets, and also utilize as a biofertilizer for rhizome production in the field. Seven endophytic bacteria were isolated from the leaf, four isolates from the leaf base, and two from the rhizome. ECS203, a gram-negative bacterium with a round shape, showed the highest N₂ fixation at 4.2 nmol C₂H₄/10⁶ cells/hr, and ECS202 showed the highest IAA synthesis at 296 μL μg ^{? 1} protein. Three selected isolates of N₂-fixing and IAA synthesizing endophytic bacteria, i.e., ECS202, ECS203, and ECS204, isolated from the leaf base, were used to reinoculate Curcuma plantlets derived from tissue culture. Then, plants were grown in sterilized sand for 2 months and weekly supplied with N-free nutrient solution. Plant growth, colonization, nitrogen fixation, and IAA synthesis were measured at two months after planting. The inoculated plants clearly showed a better performance of plant growth and yield in terms of the plant height, plant weight, leaf area, and diameter of new rhizomes compared with uninoculated plants. The chlorophyll content and N concentration of leaves and roots also increased in inoculated plants. Endophytic bacteria from inoculated plants colonized the roots, rhizome, and leaf base. Partial sequence analysis using 16S rDNA indicated that the isolate ECS202 corresponded to Sphingomonas pseudosanguinis (99.2% similarity over 1,371 bp), ECS203 to Bacillus drentensis (99.4% similarity over 1,450 bp) and ECS204 to Bacillus methylotrophicus (99.9% similarity over 13,06 bp).     

Development of thyroid gland and changes in thyroid hormone levels in Leptocephali of Japanese Eel (Anguilla japonica)

Development of the thyroid gland of the Japanese eel (Anguilla japonica) was studied with the use of tank-reared fish. A single thyroid follicle was first found in larvae at 29 days post hatching (dph), total length (TL) 12 mm. Until reaching 25 mm in TL (100 dph), leptocephali had one or two follicles per individual. The inner colloid of the follicles was weakly immuno-positive against the anti-thyroxine (T₄) antibody. The number of thyroid follicles and the immunoreactivity later increased as the larvae grew. Thyroid hormones (TH) T₄ and triiodothyronine (T₃) were not detected in premetamorphic larvae by radioimmunoassays, but became detectable during metamorphosis. The maximum level of T₄ was seen in fish at the end of metamorphosis and in just-metamorphosed juveniles, whereas T₃ reached the highest level during metamorphosis and declined toward the end of metamorphosis. The results indicated that the thyroid gland first became active during metamorphosis in the development of eel.     

Occurrence of spontaneous polyploids from the eggs obtained by artificial induction of maturation in the Japanese eel (Anguilla japonica)

The nuclear DNA contents of the Japanese eel larvae were measured to examine the genetic soundness of the eggs obtained by artificial induction of maturation and fertilization. Of the 12.4% abnormal polyploid larvae, 77.9% were triploid and the rest were haploid, tetraploid, pentaploid, or aneuploid.     

Analysis of the relationship between luminescence and toxicity of <Emphasis Type="Italic">Vibrio carchariae</Emphasis> pathogenic to shrimp

ABSTRACT:   Aquaculture deterioration is caused by Vibrio harveyi , which is one of the main shrimp pathogens. Vibrio carchariae having the same phenotypic characteristics as V. harveyi was isolated from a shrimp farm and used in this study to investigate the relationship between luminescence and toxicity. Luminescence was verified by monitoring the expression of the LuxR gene. It was found that the relationship between the expression of LuxR and toxicity was the most significant factor related to luminescence. Results showed that LuxR expression was highest after 20 h of culture, which is comparable to the highest scintillation counter luminescence value (20 h). However, toxin levels in the culture broth were highest between 24 h-36 h of culture. Significant decrease in the toxicity level was observed after 36 h. It was also found that there was a decrease in toxicity level after 12 h of incubation at room temperature of the V. carchariae culture supernatant. The effect of the addition of cell-free culture supernatant on the luminescence of V. carchariae was also determined. Results showed that the addition of cell-free supernatant from 24 h-old culture was most effective in inducing and maintaining luminescence.     

Properties of alkaline‐hydrolyzed waterfowl feather keratin

The properties of hydrolyzed feather keratin (HFK) were compared to those of hydrolyzed wool keratin (HWK) with the aim of developing better ways to utilize feather keratin waste. Amino acid analysis showed that HFK contained more hydrophobic amino acids did than HWK. Although gel permeation chromatography indicated that HFK and HWK had more low‐molecular weight peptides than their intact sources, Fourier transform infrared spectroscopy indicated that both hydrolyzed keratins retained their original secondary structure. The physical properties of HFK were evaluated by treating HFK to human hair fibers. HFK treatment enhanced significantly the surface hydrophobicity and strength of fibers, and HFK was more permeable into hair fibers. These results suggest that HFK is suitable for industrial applications to improve fibers. In addition, HFK may be suitable for raw material of products requiring both flexibility and hydrophobicity, such as films and biodegradable plastics.     

Embryological evidence identifies wing digits in birds as digits 1, 2, and 3

The identities of the digits of the avian forelimb are disputed. Whereas paleontological findings support the position that the digits correspond to digits one, two, and three, embryological evidence points to digit two, three, and four identities. By using transplantation and cell-labeling experiments, we found that the posteriormost digit in the wing does not correspond to digit four in the hindlimb; its progenitor segregates early from the zone of polarizing activity, placing it in the domain of digit three specification. We suggest that an avian-specific shift uncouples the digit anlagen from the molecular mechanisms that pattern them, resulting in the imposition of digit one, two, and three identities on the second, third, and fourth anlagens.     

Histological observation of the reproductive system in a viviparous teleost Xenotoca eiseni Rutter 1896 (Cyprinodontiformes: Goodeidae)

Xenotoca eiseni is a viviparous teleost belonging to the family Goodeidae. Here, we report histological observations of the reproductive organs in an adult male, an adult female, a pregnant female with intraovarian embryo and an extracted embryo of X. eiseni. High-resolution images of haematoxylin–eosin-stained sagittal sections revealed the detailed structure of gonads, gametes and reproductive components of the mother–embryo relationship. In the male, mature spermatozoa in the epididymis formed sperm packages. In the female, oogenesis proceeded asynchronously in the ovarian wall, and various stages of oocytes were observed in single ovary. In both sexes, genital openings were located between the anus and anal fin. Developing embryos were observed in an ovary of the pregnant female. Fine structures of components of the mother-to-embryo nutrient supply, ovarian septum and trophotaenia were observed in the pregnant ovary. An immature gonad prior to gamete formation was identified in the extracted embryo. With the aim of supporting the development and extension of studies on this viviparous teleost, we have shared our histological images as raw data in an open online archive, the 'NAGOYA repository ( http://hdl.handle.net/2237/00032456 )'. Our goal is a comprehensive understanding of the viviparous system in fish using both histological observation and molecular biology methods including genomics and proteomics.     

Intraspecific Variation of Eysarcoris guttigerus (Hemiptera: Pentatomidae) in Japanese Southwest Population Based on Mitochondrial DNA

The white-spotted globular bug Eysarcoris guttigerus (Thunberg) (Hemiptera: Pentatomidae) is widely distributed in East Asia and the Pacific region. In Japan, the species is found in grassy or composite weeds in the western area of the main islands and Ryukyu Islands of Japan. One notable characteristic of the Eysarcoris genus is the two white spots on the scutellum. This is not the case with the Ishigaki Island population, however, which sports red spots instead of white, suggesting that intraspecific variation exists in the species. Therefore, we investigated intraspecific variation in E. guttigerus using mitochondrial NADH dehydrogenase subunit 2 (ND2), cytochrome oxidase subunit 1 (CO1), cytochrome b (Cytb), tRNA-Serine (tRNA^ser), NADH dehydrogenase subunit 1 (ND1), and 16S ribosomal RNA (16SrRNA) genes from 13 populations of Japan. The obtained maximum likelihood phylogenetic tree was divided into three groups—Group 1: Mainland, Group 2: Central Ryukyu Islands (Okinawa-Amamioshima Islands), and Group 3: South Ryukyu Islands (Ishigaki Island). The Ishigaki population was significantly separated from the other populations with consistent differences in spot color. The estimated period of divergence between the Ishigaki population and the other populations was consistent with the period of formation of the Kerama Gap in the Ryukyu arc. Thus, the process of formation of the Kerama Gap may have influenced the intraspecific variation of E. guttigerus.     

Sampling method for estimating neutral allele frequency in a pedigreed population

The estimated allele frequency at neutral DNA marker loci is a fundamental parameter for establishing a conservation scheme for a set of livestock breeds. In this study, we propose a novel 'minimum distance (MD) method' for estimating neutral allele frequencies, which minimizes the error by the use of pedigree information. Using computer simulation and actual microsatellite data for a pedigreed cattle population, the performance of the proposed method was compared with that of conventional random sampling (RND). MD manifests a promising superiority to RND, irrespective of the sample size and the depth of pedigree. It is theoretically demonstrated that the error of the estimates depends on both the sample size and the distance of founder allele frequencies between the sampled and the non-sampled individuals. MD samples individuals so as to minimize the latter source of estimation error.